Despite the recent completion of the human genome project, an ostensibly more difficult post-genomic challenge will be the functional annotation of all human genes and integration of this information into a rational drug discovery process. Unfortunately, this is at present challenging, primarily due to the limited choice of suitable toolsets to rapidly delineate potential drug targets and screen new drugs en masse. Phage display is a unique tool for the discovery of peptide ligands, design of peptidomimetics, and biological validation of specific targets. Despite these successes, application of phage display technology for High Throughput Screening (HTS) of biologically active peptides in a disease-relevant cell model has not been developed so far. To address these issues, we will develop a novel experimental platform for cell-based HTS of peptide drugs as an alternative to HTS of synthetic peptide or chemical small molecule libraries. The long-term goal of the current grant application is the development and commercialization of next- generation HTS technology for the simultaneous screening of millions of peptide compounds for biological activity with pooled phage display libraries, concomitantly with a wide range of reporter cell lines compatible with this HTS technology of peptide leads. The technologically simple and cost-effective approach of cell- based HTS with pooled phage libraries displaying millions of peptide compounds, which far exceeds the complexity of small molecule libraries, will change current drug discovery strategies and accelerate the development of novel therapeutics. We propose to use our novel bioactive peptide HTS resource to delineate the processes which underlie deregulated proliferation in cancer cells, and to apply this technology to identify cytotoxic peptides specific for cancer cells. Under Phase I funding, we initially propose to develop a pooled phage library displaying peptides for 500 pharmaceutically relevant cytokines, growth factors, chemokines, and hormones, as well as technology for HTS of biologically active phage displayed peptides in a cell-based assay. In collaboration with our biology consultants at the Cleveland Clinical Foundation, we will subsequently validate this library in transcriptional reporter cell line models, to identify effector peptides of NF-?B signaling pathway. The combination of phage display technology with cell-based HTS will transform phage display technology into a powerful tool for identification and validation of novel biologically relevant drugs. Moreover, we envisage a major impact on the molecular dissection of human disease mechanisms. For example, these reagents harbor considerable promise to identify new targets for therapeutic intervention, and the development of increasingly relevant paradigms for drug discovery. As a result, we foresee that these toolsets will significantly improve the efficiency, economy, and ease of performing HTS of lead peptide compounds, and will provide both academic and industrial researchers with preferred, cost-effective alternatives to existing small molecule libraries. The ultimate goal of the proposed project is to develop and make commercially available new, powerful drug discovery tools: a technology for high throughput screening of peptide pharmaceuticals in a cell-based assay with a set of genome-wide receptor ligand phage display libraries and transcriptional reporter cell lines. We propose to validate and apply these tools for discovery peptide agonists and antagonists of NFkB signaling pathway. The developed high throughput screening tools and technologies will significantly improve the efficiency of translational research related to molecular dissection of diverse human disease mechanisms, development of new pharmaceuticals, and therefore, have major implications for improving drug discovery research. ? ? ?
